Changes in intra- and interlimb reflexes from forelimb cutaneous afferents after staggered thoracic lateral hemisections during locomotion in cats

Abstract

In quadrupeds, such as cats, cutaneous afferents from the forepaw dorsum signal external perturbations and send inputs to spinal circuits to co-ordinate the activity in muscles of all four limbs. How these cutaneous reflex pathways from forelimb afferents are reorganized after an incomplete spinal cord injury is not clear. Using a staggered thoracic lateral hemisections paradigm, we investigated changes in intralimb and interlimb reflex pathways by electrically stimulating the left and right superficial radial nerves in seven adult cats and recording reflex responses in five forelimb and ten hindlimb muscles. After the first (right T5-T6) and second (left T10-T11) hemisections, forelimb-hindlimb co-ordination was altered and weakened. After the second hemisection, cats required balance assistance to perform quadrupedal locomotion. Short-, mid- and long-latency homonymous and crossed reflex responses in forelimb muscles and their phase modulation remained largely unaffected after staggered hemisections. The occurrence of homolateral and diagonal mid- and long-latency responses in hindlimb muscles evoked with left and right superficial radial nerve stimulation was significantly reduced at the first time point after the first hemisection, but partially recovered at the second time point with left superficial radial nerve stimulation. These responses were lost or reduced after the second hemisection. When present, all reflex responses, including homolateral and diagonal, maintained their phase-dependent modulation. Therefore, our results show a considerable loss in cutaneous reflex transmission from cervical to lumbar levels after incomplete spinal cord injury, albeit with preservation of phase modulation, probably affecting functional responses to external perturbations. KEY POINTS: Cutaneous afferent inputs co-ordinate muscle activity in the four limbs during locomotion when the forepaw dorsum contacts an obstacle. Thoracic spinal cord injury disrupts communication between spinal locomotor centres located at cervical and lumbar levels, impairing balance and limb co-ordination. We investigated cutaneous reflexes from forelimb afferents during quadrupedal locomotion by electrically stimulating the superficial radial nerve bilaterally, before and after staggered lateral thoracic hemisections in cats. We showed a loss/reduction of mid- and long-latency homolateral and diagonal reflex responses in hindlimb muscles early after the first hemisection that partially recovered with left superficial radial nerve stimulation, before being reduced after the second hemisection. Targeting cutaneous reflex pathways from forelimb afferents projecting to the four limbs could help develop therapeutic approaches aimed at restoring transmission in ascending and descending spinal pathways.

Keywords: cutaneous reflexes; interlimb co‐ordination; locomotion; spinal cord injury.

Figure 1.. Experimental chronology and estimation of lesions extent.

(A) Chronology showing the first (T1) and second (T2) experimental time points after the first (H1) and second (H2) hemisections in all cats. (B) For lesions extent, the black area represents the estimation as a percentage of total cross-sectional area (reproduced with permission of (Mari et al., 2024)). Note that we only performed one lesion in cat KI.

Figure 2.. Phase-dependent modulation of cutaneous reflexes evoked in homonymous forelimb muscles during locomotion before and following staggered hemisections.

Each panel shows, from left to right, stance phases of the stimulated forelimb (empty horizontal bars) with its averaged rectified muscle activity normalized to cycle duration in the different states/time points, and homonymous reflex responses in representative cats for the left and right (A) extensor carpi ulnaris (ECU, cat KA), (B) triceps brachii (TRI, cat TO), and (C) biceps brachii (BB, cat JA). Reflex responses are shown with a post-stimulation window of 80 ms in four phases in the intact state, and after the first (H1) and second (H2) hemisections at time points 1 (T1) and/or 2 (T2). At each state/time point, evoked responses are scaled according to the largest response obtained in one of the four phases. The scale, however, differs between states/time points. Each trace is the average of 3–55 stimulated cycles. For durations of stance phases and EMG activities, we averaged 11–20 and 9–27 cycles, respectively. The dotted vertical lines in the reflex responses indicate the N1/P1, N2/P2 and N3/P3 time windows.

Figure 3.. Phase-dependent modulation of cutaneous reflexes evoked in crossed forelimb muscles during locomotion before and following staggered hemisections.

Each panel shows, from left to right, stance phases of the stimulated forelimb (empty horizontal bars) and crossed forelimb (filled horizontal bars) with its averaged rectified muscle activity normalized to cycle duration in the different states/time points, and crossed reflex responses in representative cats for the left and right (A) extensor carpi ulnaris (LECU, cat GR; RECU, cat TO), (B) triceps brachii (LTRI, cat AR; RTRI, cat JA), and (C) biceps brachii (BB, cat JA). Reflex responses are shown with a post-stimulation window of 80 ms in four phases in the intact state, and after the first (H1) and second (H2) hemisections at time points 1 (T1) and/or 2 (T2). At each state/time point, evoked responses are scaled according to the largest response obtained in one of the four phases. The scale, however, differs between states/time points. Each trace is the average of 6–64 stimulated cycles. For durations of stance phases and EMG activities, we averaged 12–20 and 7–23 cycles, respectively. The dotted vertical lines in the reflex responses indicate the N1/P1, N2/P2 and N3/P3 time windows.

Figure 4.. Phase-dependent modulation of cutaneous reflexes evoked in homolateral hindlimb muscles during locomotion before and following staggered hemisections.

Each panel shows, from left to right, stance phases of the stimulated forelimb (empty horizontal bars) and homolateral hindlimb (filled horizontal bars) with its averaged rectified muscle activity normalized to cycle duration in the different states/time points, and homolateral reflex responses in representative cats for the left and right (A) soleus (SOL, cat TO), (B) vastus lateralis (LVL, cat HO; RVL, cat TO), and (C) anterior sartorius (SRT, cat JA). Reflex responses are shown with a post-stimulation window of 80 ms in four phases in the intact state, and after the first (H1) and second (H2) hemisections at time points 1 (T1) and/or 2 (T2). At each state/time point, evoked responses are scaled according to the largest response obtained in one of the four phases. The scale, however, differs between states/time points. Each trace is the average of 6–55 stimulated cycles. For durations of stance phases and EMG activities, we averaged 10–20 and 9–23 cycles, respectively. The dotted vertical lines in the reflex responses indicate the N1/P1, N2/P2 and N3/P3 time windows.

Figure 5.. Phase-dependent modulation of cutaneous reflexes evoked in diagonal hindlimb muscles during locomotion before and following staggered hemisections.

Each panel shows, from left to right, stance phases of the stimulated forelimb (empty horizontal bars) and diagonal hindlimb (filled horizontal bars) with its averaged rectified muscle activity normalized to cycle duration in the different states/time points, and diagonal reflex responses in representative cats for the left and right (A) soleus (LSOL, cat TO; RSOL, cat JA), (B) vastus lateralis (VL, cat TO), and (C) anterior sartorius (LSRT, cat TO; RSRT, cat JA). Reflex responses are shown with a post-stimulation window of 80 ms in four phases in the intact state, and after the first (H1) and second (H2) hemisections at time points 1 (T1) and/or 2 (T2). At each state/time point, evoked responses are scaled according to the largest response obtained in one of the four phases. The scale, however, differs between states/time points. Each trace is the average of 6–55 stimulated cycles. For durations of stance phases and EMG activities, we averaged 13–20 and 8–23 cycles, respectively. The dotted vertical lines in the reflex responses indicate the N1/P1, N2/P2 and N3/P3 time windows.

Figure 6.. Reflex response occurrence in all four limbs before and after staggered hemisections.

Response occurrence probabilities are shown for short- (SLR) and mid-/long-latency (MLRs/LLRs) responses with stimulation of the left or right superficial radial nerve before (intact) and after the first (H1) and second (H2) hemisections at time points 1 (T1) and/or 2 (T2). Tables 1, 2 and 3 provide details on the number of pooled data for SLRs and MLRs/LLRs, with a N ranging from 19–30 and 19–55, respectively, at each states/time points. Each filled circle represents the mean probability ± confidence interval (95%) in 5 forelimb or 10 hindlimb muscles pooled across cats for homonymous/crossed (A and B) and homolateral/diagonal (C and D) responses, respectively. If a significant main effect of state/timepoint was found (generalized linear mixed model), we compared states/time points. Asterisks indicate significant differences at p < 0.05*, p < 0.01** and p < 0.001***. When one state/time point was significantly different from two states/time points, the comparison starts with a longer horizontal line.

Figure 7.. Schematic illustration of putative pathways and mechanisms contributing to cutaneous reflexes and their modulation before and after staggered hemisections.

In the intact state, afferents from the left (A) and right (B) superficial radial (SR) nerves contact spinal interneurons that project to motoneurons within the hemisegment (homonymous responses) at cervical levels and commissural interneurons projecting contralaterally (crossed responses). SR nerve afferents also make contacts with propriospinal neurons that project to lumbar levels, terminating ipsilaterally (homolateral responses) or contralaterally via collateral projections at different segments (diagonal responses). The pathways responsible for short-latency responses (SLRs) are mainly confined to spinal circuits, including SLRs in hindlimb muscles. The pathways contributing to mid- and long-latency responses (MLRs/LLRs) transmit sensory information to supraspinal structures via long ascending projection neurons (propriospinal and/or dorsal lemniscal pathways) that project back to spinal circuits controlling the fore- and hindlimbs. After the first hemisection (on the right side), SLRs and MLRs/LLRs in forelimb muscles remain present although their response pattern can change due to the loss of inhibitory or excitatory ascending pathways from lumbosacral circuits. The occurrence of MLRs/LLRs in hindlimb decreases (dashed lines) due to disruptions in ascending and descending pathways to and from supraspinal structures. Spared supraspinal axons are potentially strengthened or sprout to form new connections to transmit descending signals. After the second hemisection (on the left side), direct ascending and descending pathways are both disrupted, and reorganization of short propriospinal neurons is required to relay signals through the lesions, although their ability to do so is limited, leading to a considerable loss in MLRs and LLRs in hindlimb muscles.

Figure 8.. Homolateral responses in the left soleus for one cat for grouped and separated forelimb cycles.

(A) EMG activities for the left extensor carpi ulnaris (LECU) and soleus (LSOL) muscles along with the stance phases of the left forelimb (LFST) and hindlimb (LHST). We illustrate cycles where the stimulation occurred at mid-stance and mid-swing for the first and second forelimb cycles (B) Each panel shows homolateral reflex responses in four phases of the left forelimb cycle in the left soleus for cat JA, with a post-stimulation window of 80 ms. Aligned vertically on the right of each panel is the averaged rectified activity of the left soleus normalized to cycle duration of the left forelimb, along with left forelimb (red bar) and left hindlimb (grey bar) stances. Reflex responses are shown when the first and second cycles of the left forelimb are grouped together and when they are separated, after the first (H1T2) and second (H2T2) hemisections at the second time point. At each state/time point, evoked responses are scaled according to the largest response obtained in one of the four phases and the grouped/separated cycles. The scale, however, differs between states/time points. The number of averaged stimulated cycles is indicated at the left of each reflex responses.